Growth inhibition and activity stimulation of non-target organism nitrogen-fixing bacterium Azotobacter vinelandii by herbicide florasulam.

The herbicide florasulam is widely used to control broadleaf weeds in modern agriculture. Although the general ecotoxicity of herbicides has been verified, their effects on non-target soil functional bacteria, e.g., nitrogen-fixing bacteria, remained unknown. Herein, we investigated the potential impacts of florasulam on the nitrogen-fixing model bacterium Azotobacter vinelandii and sought to understand the underlying mechanisms. The growth activity of A. vinelandii was inhibited by florasulam in a concentration-dependent pattern, and the half-lethal concentration (IC50) was calculated as 8.21 mg/L. Surprisingly, the nitrogen-fixing activity and energy metabolism of A. vinelandii were stimulated by florasulam at 4-8 mg/L. Transcriptomic analyses revealed that the gene expression of nitrogen-fixing bacteria changed significantly after florasulam treatment. The genes encoding the iron nitrogenase and the key enzymes in the tricarboxylic acid cycle (TCA cycle) were significantly up-regulated. Metabolomic analyses elucidated that florasulam inhibited the folate metabolism pathways on the third and seventh days, directly impairing the DNA replication processes and consequently hindering the cell division, which led to the reduction of biotin and riboflavin levels. Notably, the up-regulation of amino acid biosynthesis, the TCA cycle, and glutathione metabolism networks likely maintained the high nitrogen fixation capacity of A. vinelandii. Therefore, florasulam enhanced nitrogen fixation and the energy metabolism in A. vinelandii through up-regulating the TCA cycle and the glutathione metabolism, although it inhibited bacterial growth.